Compound-specific protein expression signatures (PESs) can be revealed by proteomic techniques. The SELDI-TOF MS approach is advantageous due to its simplicity and high-throughput capacity, however, there are concerns regarding the reproducibility of this method. The aim of this study was to define an estrogen-responsive PES in plasma of Atlantic cod (Gadus morhua) using the SELDI-TOF MS technique. Protein expression analysis of male cod exposed to 17β-estradiol (E₂) showed that 27 plasma peaks were differentially expressed following exposure. The reproducibility of this result was evaluated by reanalyzing the samples six months later, and a significant change in expression was confirmed for 13 of the 27 peaks detected in the first analysis. The performance of the reproducible E₂-responsive PES, constituting these 13 peaks, was then tested on samples from juvenile cod exposed to 4-nonylphenol, North Sea oil, or North Sea oil spiked with alkylphenols. Principal component analysis revealed that nonylphenol-exposed cod could be separated from unexposed cod based on the E₂-responsive PES, indicating that the PES can be used to assess estrogenic exposure of both juvenile and adult specimens of cod. A targeted antibody-assisted SELDI-TOF MS approach was carried out in an attempt to identify the E₂-responsive peaks. Results indicated that 2 peaks were fragments of the well-known biomarkers VTG and/or ZRP. In this study, the SELDI-TOF MS technology has shown its potential for defining compound-specific PESs in fish. Nevertheless, thorough validation of reproducibility, specificity and sensitivity of a PES is required before it can be applied in environmental monitoring.
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